Correlation of calcitonin structure to biological activity has proven difficult. Often the combination of single amino acid substitutions (ability to reduce level of serum calcium) has not resulted in additive effects on biological activity, perhaps due to a lack of information on the metabolic properties of the hormone. There is a wide variation in activity in the naturally occurring calcitonins with an approximate 40-fold range in biopotency. All calcitonins share some common structural features. Each is 32 amino acids long with a C-terminal prolinamide and an N-terminal disulfide linked ring from position 1 through 7. Salmon 1 calcitonin, for example, has the following formula (Niall, H. D. (1969) Proc. Natl. Acad. Sci. USA 64, 771-778): ##STR1## Other natural calcitonins have different degrees of homology with this sequence (Queener, S. F. and Bell, N. H. (1975) Metabolism 24, 555-567; Lasmoles, F. et al. (1985) FEBS Lett. 180, 113-116). The amino acid L-threonine (2S,3R configuration) at position 31 also occurs in eel, ovine, bovine, porcine and chicken calcitonins. The same position 31 is occupied by the amino acid L-alanine in human, murine and L-valine in salmon 2 calcitonin and salmon 3 calcitonin. Eel calcitonin differs from salmon 1 calcitonin by having the amino acids L-Asp at position 26, L-Val at position 27 and L-Ala at position 29.
Chicken calcitonin differs from salmon 1 calcitonin by having the amino acid L-Ala at position 2, L-Ser at position 3, L-Asp at position 26, L-Val at position 27 and L-Ala at position 29. Salmon 2 calcitonin differs from salmon 1 calcitonin by having L-Asp at position 15, L-Phe at position 22, L-Ala at position 29 and L-Val at position 31. Salmon 3 calcitonin differs from salmon 1 calcitonin by having L-Met at position 8, L-Asp at position 15, L-Phe at position 22, L-Ala at position 29 and L-Val at position 31.
The calcitonins of mammalian origin differ more markedly from salmon 1 calcitonin, as shown by the following comparison. The disulfide linkage between positions 1 and 7 is omitted from the following sequence for clarity.
______________________________________ Position: 1 2 3 4 5 6 7 ______________________________________ Salmon 1 Cys Ser Asn Leu Ser Thr Cys Human " Gly " " " " " Murine " " " " " " " Bovine " Ser " " " " " Porcine " " " " " " " Ovine " " " " " " " ______________________________________ Position: 8 9 10 11 12 13 14 ______________________________________ Salmon 1 Val Leu Gly Lys Leu Ser Gln Human Met " " Thr Tyr Thr " Murine " " " " " " " Bovine Val " Ser Ala " Trp Lys Porcine " " " " " " Arg Ovine " " " " " " Lys ______________________________________ Position: 15 16 17 18 19 20 21 ______________________________________ Salmon 1 Glu Leu His Lys Leu Gln Thr Human Asp Phe Asn " Phe His " Murine " Leu " " " " " Bovine " " " Asn Tyr " Arg Porcine Asn " " " Phe " " Ovine Asp " " " Tyr " " ______________________________________ Position: 22 23 24 25 26 27 28 ______________________________________ Salmon 1 Tyr Pro Arg Thr Asn Thr Gly Human Phe " Gln " Ala Ile " Murine " " " " Ser " " Bovine " Ser Gly Met Gly Phe " Porcine " " " " " " " Ovine Tyr " " " " " " ______________________________________ Position: 29 30 31 32 ______________________________________ Salmon 1 Ser Gly Thr Pro--NH.sub.2 Human Val " Ala " Murine " " " " Bovine Pro Glu Thr " Porcine " " " " Ovine " " " " ______________________________________
The structural features responsible for the increased potency of ultimobranchial calcitonins relative to calcitonins of mammalian origin have not yet been fully determined. The removal of the C-terminal amide group to a proline moiety in human calcitonin results in a drastic loss of the activity, from 100% to 0.06%. The deletion of the entire prolinamide moiety also gives a drastic loss in activity, from 100% to 0.12% (Rittel, W. et al. (1976) Experientia 32, 246-248).
U.S. Pat. No. 4,469,632 shows the replacement of naturally occurring Arg.sup.24 with D-Arg.sup.24 in salmon 1 calcitonin, with increased activity. On the other hand, U.S. Pat. No. 4,414,149 shows a reduction in activity of salmon 1 calcitonin when L-Val.sup.8 and L-Arg.sup.24 are respectively replaced with L-Gly.sup.8 and D-Arg.sup.24.